The main focus of this research proposal is to determine the molecular mechanisms by which spontaneous neurotransmitter release is regulated. Characterization of neuronal communication at synapses has largely focused on action potential-triggered synaptic vesicle fusion, with spontaneous miniature potentials (minis) largely thought to represent background noise. However, spontaneous release is regulated independently of evoked release. Moreover, the frequency of spontaneous release is regulated by activity and can drive synaptic structural modification and growth, changes resulting in long-lasting alterations in neuronal connectivity. Regulation of spontaneous release would likely impinge directly on the molecular "fusion clamp" which prevents primed vesicles from fusing with the presynaptic membrane in the absence of calcium, Complexin has been identified as the vesicle fusion clamp that regulates spontaneous release at the Drosophila neuromuscular junction (NMJ). However, the molecular mechanism by which complexin functions as a fusion clamp to regulate spontaneous release is unknown. The Drosophila NMJ will be used as an in vivo model system to determine the molecular mechanisms by which complexin regulates spontaneous neurotransmitter release. Aim 1 will examine evolutionary conservation of complexin function as a vesicle fusion clamp. Aim 2 will define the molecular mechanisms that mediate complexin's ability to regulate spontaneous release. Both aims will make extensive use of genetic tools available in Drosophila and in vivo electrophysiology recordings, as well as in vitro biochemical and immunocytochemical approaches. Alterations in complexin levels have been reported in a number of neurological diseases including schizophrenia, Huntington's disease, and Alzheimer's disease, suggesting the complexin dysfunction and abnormal rates of spontaneous release may contribute to several human neuropathologies. Defects in neurotransmitter release are implicated in a number of neurological diseases including schizophrenia, Huntington's disease, and Alzheimer's disease. Understanding the molecular mechanisms that underlie neurotransmitter release will provide potentially new targets and insights into how altered spontaneous release contributes to neurological diseases.